Pan-azole-resistant Candida tropicalis carrying homozygous erg11 mutations at position K143R: a new emerging superbug?

Objectives: Candidaemia is a public health problem mainly in hospitalized individuals worldwide. In Brazil, Candida albicans is the most prevalent species that causes candidaemia, followed by Candida tropicalis and Candida parapsilosis . Few data on the abundance of antifungal resistance are available for Latin America. Methods: We analysed the frequency of azole and echinocandin resistance in Candida isolates ( n = 75) collected between 2012 and 2014 at the University Hospital of Federal University of Juiz de Fora (Brazil). The primary targets erg11 (azoles) and fks1 (echinocandins) were sequenced and modelled at the protein level. Antifungal susceptibility testing was performed according to CLSI (M27-A3 and M27-S4) and according to EUCAST. Results: The three most frequent species were C. albicans (38.0%), C. tropicalis (30.0%) and Candida glabrata (17.0%). Azole resistance was observed in 27.0% of all Candida isolates, while 20.0% of all isolates were echinocandin resistant. A novel mutation in erg11 at location K143R was found to be associated with phenotypically pan-azole-resistant C. tropicalis isolates. This mutation maps near the active binding site of erg11 and is likely to confer pan-azole resistance to C. tropicalis . Conclusions: A novel point mutation (K143R) located in the erg11 gene of C. tropicalis was found in pan-azole-resistant strains. According to our protein homology model, it is very likely that the mutation K143R causes pan-azole resistance in C. tropicalis . Moreover, an up-regulation of ABC transporters was observed, which can add up to a pan-azole-resistant phenotype.

Peptidorhamnomannan negatively modulates the immune response in a scedosporiosis murine model.

In this study, we analyzed the impact of immunization with the peptidorhamnomannan (PRM) from the cell wall of the fungus Scedosporium (Lomentospora) prolificans in a murine model of invasive scedosporiosis. Immunization with PRM decreased the survival of mice infected with S. prolificans. Immunization of mice with PRM led to decreased secretion of pro-inflammatory cytokines and chemokines but did not affect the secretion of IL-10. Mice immunized with PRM showed an increase in IgG1 secretion, which is an immunoglobulin linked to a nonprotective response. Splenocytes isolated from mice infected with S. prolificans and immunized with PRM showed no differences in the percentages of Th17 cells and no increase in the frequency of the CD4(+)CD62L(Low) T cell population. PRM-immunized mice showed a significant increase in the percentage of Treg cells. In summary, our results indicated that immunization with PRM did not assist or improve the immunological response against S. prolificans infection. PRM exacerbated the infection process by reducing the inflammatory response, thereby facilitating colonization, virulence and dissemination by the fungus.

O-glycosylation in cell wall proteins in Scedosporium prolificans is critical for phagocytosis and inflammatory cytokines production by macrophages.

In this study, we analyze the importance of O-linked oligosaccharides present in peptidorhamnomannan (PRM) from the cell wall of the fungus Scedosporium prolificans for recognition and phagocytosis of conidia by macrophages. Adding PRM led to a dose-dependent inhibition of conidia phagocytosis, whereas de-O-glycosylated PRM did not show any effect. PRM induced the release of macrophage-derived antimicrobial compounds. However, O-linked oligosaccharides do not appear to be required for such induction. The effect of PRM on conidia-induced macrophage killing was examined using latex beads coated with PRM or de-O-glycosylated PRM. A decrease in macrophage viability similar to that caused by conidia was detected. However, macrophage killing was unaffected when beads coated with de-O-glycosylated PRM were used, indicating the toxic effect of O-linked oligosaccharides on macrophages. In addition, PRM triggered TNF-α release by macrophages. Chemical removal of O-linked oligosaccharides from PRM abolished cytokine induction, suggesting that the O-linked oligosaccharidic chains are important moieties involved in inflammatory responses through the induction of TNF-α secretion. In summary, we show that O-glycosylation plays a role in the recognition and uptake of S. prolificans by macrophages, killing of macrophages and production of pro- inflammatory cytokines.